Chromosome replication and transmission are essential for the inheritance of genetic traits, but the mechanisms responsible for these processes remain poorly understood in multicellular eukaryotes. The centromere, which appears as a constriction in metaphase chromosomes, is associated with the kinetochore and serves as the key attachment site to the spindle during mitosis and meiosis. A pressing question in the centromere field today is how centromere identity is propagated from one generation to the next in multicellular eukaryotes. Elucidating the determinants of centromere identity, propagation and function requires identification of the gene products that promote centromere formation and function in trans, and ultimately determining how they interact with the cis-acting elements to perform their essential functions. In essence, we need to understand the organization, replication, and modification of centromeric chromatin. The multifaceted genetic, molecular, cell biological and biochemical approaches required to address these complex questions in higher eukaryotes are likely to succeed in Drosophila. There is over a century of experimental analyses using the fruit fly, which facilitates sophisticated in vivo analyses. Here, we propose genetic, molecular, cell biological and biochemical experiments designed to identify and characterize gene products that promote the assembly and propagation of centromeric chromatin, and to determine their properties and functions. Our entry point into centromeric chromatin will be a conserved histone H3-1ike protein (CID, for Centromere IDentifier) that localizes exclusively to functional centromeres. We will use genetic screens and biochemical isolations to identify genes and proteins that interact with CID, then use a variety of approaches to determine their roles in centromere function. These studies will address specific hypotheses and provide the groundwork for future analysis of inheritance and centromere function in Drosophila and other higher eukaryotes, such as humans.